Pretreatment of pituitary cell cultures with GnRH causes altered gonadotrope responsiveness to LH secretagogues. The precise mechanism by which this occurs is not understood. Because a G protein appears to be activated after GnRH stimulation of the gonadotrope, a role for this moiety in GnRH-stimulated alterations in gonadotrope responsiveness was assessed. We show that 3 h pretreatment of pituitary cell cultures with 10 mM NaF (a G protein activator), resulted in decreased gonadotrope responsiveness to subsequent GnRH treatment (3 h, 100 nM; 34.4 +/- 1.6% vs. 23.4 +/- 1.5% of total cellular LH). NaF-provoked gonadotrope desensitization to GnRH also occurred in the presence of 3 mM EGTA and in cells which had been depleted of protein kinase C. Desensitization to GnRH did not occur in response to pretreatment with (Bu)2cAMP (8 h, 1 mM). In addition, neither GnRH nor NaF stimulated inositol phosphate production above basal levels after the NaF pretreatment. GnRH receptor binding also decreased by 30% with NaF pretreatment. In contrast, 3 h NaF (10 mM) pretreatment enhanced responsiveness of the gonadotrope to the Ca2+ ionophore A23187 in a protein kinase C- and cAMP-dependent manner. Responsiveness to the phorbol ester, phorbol 12-myristate 13-acetate, was also increased, whereas responsiveness to the Ca2+ channel activator maitotoxin was unchanged. These data suggest that G protein activation by NaF provokes gonadotrope desensitization to GnRH stimulation by both decreasing receptor numbers and by uncoupling of the receptors from inositol phosphate production. In addition, a distinct G protein action appears to be involved in sensitizing the gonadotrope to A23187 and phorbol 12-myristate 13-acetate.